PIAS1 mediates TGFbeta-induced SM alpha-actin gene expression through inhibition of KLF4 function-expression by protein sumoylation.
OBJECTIVE
TGFbeta and proliferation/phenotypic switching of smooth muscle cells (SMCs) play a pivotal role in pathogenesis of atherosclerotic and restenotic lesions after angioplasty. We have previously shown that the protein inhibitor of activated STAT (PIAS)1 activates expression of SMC differentiation marker genes including smooth muscle (SM) alpha-actin by interacting with serum response factor (SRF) and class I bHLH proteins. Here, we tested the hypothesis that TGFbeta activates SM alpha-actin through PIAS1.


RESULTS
An siRNA specific for PIAS1 and ubc9, an E2-ligase for sumoylation, inhibited TGFbeta-induced expression of SM alpha-actin in cultured SMCs as determined by real-time RT-PCR. Overexpression of PIAS1 increased SM alpha-actin promoter activity in a TGFbeta control element (TCE)-dependent manner. Because the TCE within the SM alpha-actin promoter could mediate repression through interaction with KLF4, we tested whether PIAS1 regulates the function of KLF4 for SMC gene expression. PIAS1 interacted with KLF4 in mammalian two-hybrid and coimmunoprecipitation assays, and overexpression of PIAS1 inhibited KLF4-repression of SM alpha-actin promoter activity. Moreover, PIAS1 promoted degradation of KLF4 through sumoylation.


CONCLUSIONS
These results provide evidence that PIAS1 promotes TGFbeta-induced activation of SM alpha-actin gene expression at least in part by promoting sumoylation and degradation of the TCE repressor protein, KLF4.
Kawai-Kowase
Keiko
K
Department of Medicine and Biological Science, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma, 371-8511, Japan.
Ohshima
Takayuki
T
Matsui
Hiroki
H
Tanaka
Toru
T
Shimizu
Takehisa
T
Iso
Tatsuya
T
Arai
Masashi
M
Owens
Gary K
GK
Kurabayashi
Masahiko
M
D000199:Actins / Q000235:genetics*; D000818:Animals; D001011:Aorta / Q000502:physiology; D019556:COS Cells / Q000166:cytology / Q000502:physiology; D002522:Chlorocebus aethiops; D005786:Gene Expression Regulation / Q000187:drug effects*; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000037:antagonists & inhibitors* / Q000502:physiology; D051379:Mice; D008807:Mice, Inbred BALB C; D009131:Muscle, Smooth, Vascular / Q000502:physiology; D050828:Protein Inhibitors of Activated STAT / Q000502:physiology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D000199:Actins; C000715468:Klf4 protein, mouse; C000715469:Klf4 protein, rat; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; C494211:Pias1 protein, mouse; D050828:Protein Inhibitors of Activated STAT; D034741:RNA, Small Interfering; D016212:Transforming Growth Factor beta; D012313:RNA